Fractionation-free negative enriching for in-depth C-terminome analysis

作     者：Jingtian Lu Ting Wang Huimin Bao Haojie Lu Jingtian Lu;Ting Wang;Huimin Bao;Haojie Lu

作者机构：Department of Chemistry and Shanghai Cancer CenterFudan UniversityShanghai 200438China Institutes of Biomedical Sciences and NHC Key Laboratory of Glycoconjugates ResearchFudan UniversityShanghai 200032China 

出 版 物：《Chinese Chemical Letters》 (中国化学快报（英文版）)

年 卷 期：2022年第33卷第3期

页      面：1343-1345页

核心收录：

学科分类：0710[理学-生物学] 071010[理学-生物化学与分子生物学] 07[理学] 0703[理学-化学] 

基　　金：supported by the National Key Research and Development Program of China (No. 2017YFA0505001) National Natural Science Foundation of China (No. 21974025) the project of Shanghai Key Laboratory of Kidney and Blood Purification。 

主　　题：C-terminome In-depth analysis Negative enrichment Neo-C-termini Methylamidation HeLa 

摘      要：Herein, we developed a fractionation-free negative enriching method incorporating methylamidation, siteselective dimethylation and aldehyde resin coupling(MADMAR) for in-depth C-terminome analysis. The methylamidation blocked the free carboxyl group on proteins first, followed by Lys C digestion of methylamidated proteins. Then, the site-selective dimethylation blocked the N-terminal amino group of the digested peptides without affecting the amino groups of lysine. Finally, the aldehyde resin was used to capture non-C-terminal peptides containing amino groups from lysine, while leaving the C-terminal peptides without free amino group in the supernatant for its analysis. We identified 1359 database-annotated protein C-termini from 50 μg He La proteins, which was 74% more than our previous method based on aldehyde resin. Moreover, 279 protein neo-C-termini were identified.